S Chatterjee1, I Levitan, Z Wei, A B Fisher. 1. Institute for Environmental Medicine, University of Pennsylvania, School of Medicine, Philadelphia, Pennsylvania 19104-6068, USA.
Abstract
OBJECTIVE: To investigate the role of a KATP channel in sensing shear, specifically its cessation, in the endothelial cells of the pulmonary microvasculature. METHODS: Endothelial cells isolated from the pulmonary microvasculature of wild-type and KATP channel knockout (KIR6.2-/-) mice were either statically cultured (non-flow-adapted) or kept under flow (flow-adapted) and the KIR currents in these cells were monitored by whole-cell patch-clamp technique during flow and its cessation. Membrane potential changes, generation of reactive oxygen species (ROS), and Ca2+ influx with flow cessation were evaluated by the use of fluorescent dyes. Lungs isolated from wild-type mice were imaged to visualize ROS generation in the subpleural endothelium. RESULTS: By patch-clamp analysis, reduction in the KIR current with cessation of flow occurred only in wild-type cells that were flow-adapted and not in flow-adapted KIR6.2-/- cells. Similar observations were made using changes in bisoxonol fluorescence as an index of cell membrane potential. Generation of ROS and Ca2+ influx that follow membrane depolarization were significantly lower in statically cultured and in KIR6.2-/- cells as compared to flow-adapted wild-type cells. Imaging of subpleural endothelial cells of the whole lung showed that the KATP antagonist glyburide caused the production of ROS in the absence of flow cessation. CONCLUSIONS: The responses to stop of flow (viz. membrane depolarization, KIR currents, ROS, Ca2+) were significantly altered with knockout of KATP channels, which indicates that this channel is an important component of the pulmonary endothelial response to abrupt loss of shear stress.
OBJECTIVE: To investigate the role of a KATP channel in sensing shear, specifically its cessation, in the endothelial cells of the pulmonary microvasculature. METHODS: Endothelial cells isolated from the pulmonary microvasculature of wild-type and KATP channel knockout (KIR6.2-/-) mice were either statically cultured (non-flow-adapted) or kept under flow (flow-adapted) and the KIR currents in these cells were monitored by whole-cell patch-clamp technique during flow and its cessation. Membrane potential changes, generation of reactive oxygen species (ROS), and Ca2+ influx with flow cessation were evaluated by the use of fluorescent dyes. Lungs isolated from wild-type mice were imaged to visualize ROS generation in the subpleural endothelium. RESULTS: By patch-clamp analysis, reduction in the KIR current with cessation of flow occurred only in wild-type cells that were flow-adapted and not in flow-adapted KIR6.2-/- cells. Similar observations were made using changes in bisoxonol fluorescence as an index of cell membrane potential. Generation of ROS and Ca2+ influx that follow membrane depolarization were significantly lower in statically cultured and in KIR6.2-/- cells as compared to flow-adapted wild-type cells. Imaging of subpleural endothelial cells of the whole lung showed that the KATP antagonist glyburide caused the production of ROS in the absence of flow cessation. CONCLUSIONS: The responses to stop of flow (viz. membrane depolarization, KIR currents, ROS, Ca2+) were significantly altered with knockout of KATP channels, which indicates that this channel is an important component of the pulmonary endothelial response to abrupt loss of shear stress.
Authors: James C Lee; Paul A Kinniry; Evguenia Arguiri; Matthew Serota; Stathis Kanterakis; Shampa Chatterjee; Charalambos C Solomides; Prashanthi Javvadi; Constantinos Koumenis; Keith A Cengel; Melpo Christofidou-Solomidou Journal: Radiat Res Date: 2010-05 Impact factor: 2.841
Authors: James C Lee; Ryan Krochak; Aaron Blouin; Stathis Kanterakis; Shampa Chatterjee; Evguenia Arguiri; Anil Vachani; Charalambos C Solomides; Keith A Cengel; Melpo Christofidou-Solomidou Journal: Cancer Biol Ther Date: 2009-01-01 Impact factor: 4.742